Richard W. Gray

The calciuria of increased fixed acid production in humans: Evidence against a role for parathyroid hormone and 1,25(OH)2-vitamin D

SummaryWe measured mineral and acid balances, serum iPTH, urinary cAMP/creatinine, and plasma concentrations of 25OHD and 1,25(OH)2D in 7 healthy adults during control conditions and during increased fixed acid production achieved either by the administration of NH4Cl (N=3) or by increased dietary protein intake (N=4). When acid production was increased, the subjects were in positive acid balance and negative Ca balance because of increased urinary Ca excretion. Serum iPTH fell slightly but urinary cAMP and the plasma levels of vitamin D metabolites did not change. We conclude that the accelerated skeletal and urinary losses of Ca that occur when fixed acid production is increased are not contributed to nor compensated for by the parathyroid-vitamin D endocrine systems.

Hypercalciuria and Stones

Hypercalciuria, defined as the urinary excretion of more than 0.1 mmol Ca/kg/d (4 mg/kg/24 h), is observed in approximately 50% of patients with calcium oxalate/apatite nephrolithiasis and is one of the risk factors for stone formation. Urinary Ca excretion rates among such patients are higher than normal, despite comparable ranges of glomerular filtration rate (GFR) and serum ultrafiltrable Ca concentrations, and thus glomerular filtration of Ca, suggesting that hypercalciuria is the result of inhibition of net tubular Ca reabsorption. Although increased dietary NaCl or protein intake and reduced K intake increase urinary Ca excretion rates, urinary Ca excretion rates are higher among hypercalciuric stone formers than among normal subjects in relation to comparable ranges of urinary Na, SO4 (as a reflection of protein intake), or K excretion rates, indicating that these dietary factors are not primarily responsible for hypercalciuria. Hypophosphatemia is observed among a subset of hypercalciuric patients and consequent activation of 1,25-(OH)2-D synthesis increases intestinal Ca absorption and urinary calcium excretion. Other hypercalciuric patients exhibit augmented intestinal Ca absorption without elevated plasma 1,25-(OH)-2-D levels, suggesting that either the capacity of 1,25-(OH)2-D to upregulate its own receptor in the intestine or 1,25-(OH)2-D-independent intestinal Ca transport are responsible for increased Ca absorption and hypercalciuria. Hypercalciuric patients also exhibit accelerated radiocalcium turnover, negative Ca balances, reduced bone density, delayed bone mineralization, fasting hypercalciuria, and increased hydroxyproline excretion, all of which reflect participation of the skeleton and presumably a more generalized acceleration of Ca transport. Hypercalciuria may be familial.(ABSTRACT TRUNCATED AT 250 WORDS)

Picric acid methods greatly overestimate serum creatinine in mice: More accurate results with high-performance liquid chromatography☆

The creatinine levels of blood and urine from humans, rats, and mice were measured by high-performance liquid chromatography. These were compared to the alkaline picrate analysis of creatinine performed by standard colorimetric, kinetic, and AutoAnalyzer techniques. For human serum and urine the values obtained using the HPLC technique gave good agreement with four out of five alkaline picrate techniques. For black or white mice, the serum creatinine concentration was 8.7 +/- 0.4 microM by HPLC but 44.9 +/- 1.9 microM by the lowest alkaline picrate method. Mouse urine creatinine concentrations were 3.24 +/- 0.19 mM by HPLC and 4.59 +/- 0.39 mM by the nearest alkaline picrate method. Rat serum creatinine concentrations analyzed by HPLC were about half the values obtained by AutoAnalyzer. Mouse and rat samples seemed to have substances which gave nonspecific color and thus interfered with the analysis of creatinine by the alkaline picrate methods. While the alkaline picrate analysis of creatinine was adequate for human samples, it was necessary to use HPLC to accurately measure rodent creatinine. The fractional excretion of creatinine was determined by measuring creatinine in mouse urine and plasma by both the kinetic and HPLC methods and comparing these values to urine and plasma inulin. Using the kinetic method, creatinine was cleared at 43 +/- 3% of the rate of inulin. Using the HPLC method, creatinine was cleared at 170 +/- 11% of the rate of inulin.

Impaired 24,25-Dihydroxyvitamin D Production in Anephric Human and Pig

Plasma 25-hydroxyvitamin D and 24, 25-dihydroxy-vitamin D [24,25-(OH)(2)D] concentrations were measured in normal and chronically dialyzed anephric humans and pigs. Measurement of the 24, 25-(OH)(2)D was preceded by three purification steps involving one Sephadex LH-20 column and two high-pressure liquid chromatographic columns. The final high-pressure liquid chromatography step involved resolution of 25-hydroxy-vitamin D(3)-26,23 lactone and 25,26-dihydroxy-vitamin D(2) from 24,25-dihydroxyvitamin D(2) and 24,25-dihydroxyvitamin D(3) [24,25-(OH)(2)D(3)]. The total 25-hydroxyvitamin D [25-hydroxyvitamin D(2) plus 25-hydroxyvitamin D(3) (25-OHD(3))] was 31.7+/-3.6 ng/ml in the plasma of eight anephric human subjects and 40.1+/-3.7 ng/ml in five normal human subjects. Six of the eight anephric patients had undetectable (<0.2 ng/ml) 24,25-(OH)(2)D concentrations. Two of the eight patients had very low (0.51 and 0.41 ng/ml), but detectable, 24,25-dihydroxyvitamin D(2). The normal human volunteers had plasma 24,25-(OH)(2)D concentrations of 2.8+/-0.7 ng/ml. Chronically dialyzed anephric and normal pigs were given intramuscular injections of massive amounts (5 x 10(6) IU) of vitamin D(3) immediately after surgery (day 0) and again on day 7. In anephric pigs, plasma 25-OHD(3) progressively rose from 12+/-4 ng/ml on day 0 to 705+/-62 ng/ml on day 10. The 25-OHD(3) concentrations in normal pigs rose from 8+/-2 ng/ml on day 0 to 439+/-64 ng/ml on day 10. Plasma 25-OHD(3) was higher in anephrics throughout the experiment, and concentrations were significantly higher (P < 0.05) on days 9 and 10. Plasma 24,25-(OH)(2)D(3) concentrations declined progressively in anephric pigs from 3.6+/-0.6 ng/ml on day 0 to 3.2+/-0.7 ng/ml on day 2. During days 4-10, plasma 24,25-(OH)(2)D(3) was not apparent until plasma 25-OHD(3) was >400 ng/ml. In control pigs, plasma 24,25-(OH)(2)D(3) was elevated from 4.3+/-0.6 ng/ml on day 0 to 178+/-2.7 ng/ml on day 3. Plasma 24,25-(OH)(2)D(3) was significantly higher (P < 0.05) in controls on days 1-8. At the end of the experiment (day 10), 24,25-(OH)(2)D(3) concentrations were similar and not significantly different in both groups (87.0+/-18.4 ng/ml in anephric and 110.3+/-32.1 ng/ml in normal pigs). The identity of the 24,25-(OH)(2)D(3) isolated from anephric pig plasma was confirmed by mass spectroscopy. Our data suggest that anephric humans receiving normal dietary levels of vitamin D(3) have little or no ability to produce 24,25-(OH)(2)D. However, we have shown that pigs produce 24,25-(OH)(2)D(3) when plasma 25-OHD(3) is extremely high (>400 ng/ml).

Effects of age and sex on the regulation of plasma 1,25-(OH)2-D by phosphorus in the rat

SummaryDietary phosphate deprivation in women, but not men, is accompanied by a fall in plasma PO4 and a rise in plasma 1,25-(OH)2-vitamin D concentrations. In contrast, young male rats exhibit a fall in plasma PO4 and a rise in plasma 1,25-(OH)2-D concentrations in response to PO4 deprivation. To evaluate whether age and sex influence basal plasma 1,25-(OH)2-D levels and their regulation by PO4 deprivation, plasma 1,25-(OH)2-D, PO4, and Ca levels were measured in male and female rats ranging in age from 6 weeks to 6 months while they were eating normal or low PO4 diets for 1 to 16 days. Similar observations were also made in 6-week-old castrated male and female rats, males replaced with testosterone, and females replaced with estradiol. Basal plasma 1,25-(OH)2-D levels were higher in 6-week-old males (228±76 pmol/l) than in 6-week-old females (148±62 pmol/l;P<0.01) and declined by age 11 weeks to stable levels averaging about 100 pmol/l without sex difference. Dietary PO4 deprivation resulted in a three-to fourfold increase in plasma 1,25-(OH)2-D concentrations regardless of age and sex, accompanied by a correlated rise in serum Ca concentrations. Castration of 6-week-old males and females eliminated the sex difference in basal plasma 1,25-(OH)2-D levels and appeared to enhance the elevation of plasma 1,25-(OH)2-D concentrations in response to PO4 deprivation in females. Although gonadal hormones may modify basal plasma 1,25-(OH)2-D levels, they are not required for the augmentation of plasma 1,25-(OH)2-D levels in response to PO4 deprivation.

Growth hormone and triiodothyronine permit an increase in plasma 1,25(OH)2D concentrations in response to dietary phosphate deprivation in hypophysectomized rats

SummaryHypophysectomy abolishes the four- to fivefold increase in plasma 1,25(OH)2D levels that normally accompanies dietary phosphate deprivation in rats despite a smaller but significant decrease in plasma phosphate in these animals. This effect appears within 1 week of hypophysectomy and may be the result of a lack of GH, T3, or some other pituitary hormone. In hypothyroid rats (2 weeks after TPTX) not given replacement T3, plasma 1,25(OH)2D levels rose threefold from 148±57 pmol/l to 402±96 pmol/l (mean±SD) after 4 days of dietary phosphate deprivation. However, in hypophysectomized animals given replacement T3 alone, plasma 1,25(OH)2D levels rose fourfold from 82±13 to 333±230 pmol/l after 4 days of phosphate deprivation. In addition, in hypophysectomized animals replaced with GH alone, plasma 1,25(OH)2D levels rose from 243±86 to 525±85 pmol/l during phosphate deprivation. These results would suggest that both GH and T3 must be absent to prevent enhanced renal 1,25(OH)2D synthesis during phosphate deprivation. GH and T3 appear to play a permissive role since plasma levels of these hormones do not increase when intact rats are deprived of phosphate. Furthermore, bioassayable somatomedin levels are also not increased in intact rats during phosphate deprivation as well as plasma levels of prolactin. As observed previously, plasma 1,25(OH)2D levels were inversely correlated to plasma phosphate concentrations (r=0.46,P<0.025), despite the inclusion of data points for unreplaced hypophysectomized animals who were hypophosphatemic but showed no increase in plasma 1,25(OH)2D. Thus the possibility remains that GH and T3 may exert their effect by permitting the renal 25OHD-1α-hydroxylase to respond to a change in phosphate concentrations during dietary phosphate deprivation, that, in turn, may ultimately increase renal 1,25(OH)2D synthesis and plasma levels of this hormone.

Control of plasma 1,25-(OH)2-vitamin D concentrations by calcium and phosphorus in the rat: Effects of hypophysectomy

SummaryThe mechanism by which dietary phosphate deprivation elevates plasma 1,25-(OH)2-D levels is not known. To evaluate the role of the pituitary in regulating plasma 1,25-(OH)2-D concentrations, the responses of plasma 1,25-(OH)2-D to dietary phosphate deprivation and, separately, to dietary calcium deprivation were evaluated in intact and hypophysectomized male rats. Among intact and hypophysectomized rats eating normal diets, plasma 1,25-(OH)2-D levels averaged 228±76 and 148±62 pmol/1, respectively (P<0.01). During dietary phosphate deprivation, plasma 1,25-(OH)2-D levels rose to 1160±260 in intact rats and fell to 90±26 pmol/l in hypophysectomized rats (P<0.001). By contrast, during dietary calcium deprivation, plasma 1,25-(OH)2-D levels rose in both intact and hypophysectomized animals to 856±107 and 742±279 pmol/l, respectively (NS). In response to dietary phosphate deprivation, serum calcium concentrations rose as 1,25-(OH)2-D concentrations rose in intact rats but remained at control levels in hypophysectomized rats. These results support the hypothesis that a pituitary hormone acting either directly or indirectly on the kidney mediates the increase in plasma 1,25-(OH)2-D during dietary phosphate deprivation. The hypercalcemia that occurs in rats during dietary phosphate deprivation appears to depend on the elevation of plasma 1,25-(OH)2-D.

Calcitriol, Calcium, and Granulomatous Disease

Hypercalcemia is known to occur among patients with several granulomatous diseases, including sarcoidosis,1 tuberculosis,2 berylliosis,3 and coccidioidomycosis.4 Hypercalciuria may occur even more ...

Effect of age on plasma 1,25-(OH)2 vitamin D in the rat

Levels of plasma calcium, phosphorus, creatinine and 1,25-(OH)2-D were measured in healthy male Sprague-Dawley rats aged 6 months (mature) and 20–24 months (senescent). Plasma 1,25-(OH)2-D levels fell from 95 ± 50 pmol/l in the mature rats to 41 ± 10 pmol/l in the senescent rats (p < 0.01). Despite a significant fall in plasma phosphate from 3.06 ± 0.37 mmol/l to 2.54 mmol/l (p < 0.005) in the two groups, respectively, plasma calcium remained constant at about 2.4 mmol/l in both groups.

Effects of weight loss on serum 1,25-(OH)2-vitamin D concentrations in adults: A preliminary report

SummaryDuring a review of 42 metabolic studies in healthy women and men we observed that serum 1,25-(OH)2-D concentrations were directly correlated to the observed daily changes in body weight (r=0.68;P<0.001) and to caloric intake/kg/day (r=0.39;P=0.01). These relationships could not be accounted for by related and physiologically expected changes in serum Ca or iPTH concentrations. However, serum 1,25-(OH)2-D concentrations were observed to be inversely correlated to serum PO4 levels (r=−0.44;P=0.004). In addition, serum PO4 levels were inversely correlated to the daily changes in body weight (r=−0.40;P=0.009). Since dietary sodium intake averaged 142 mmol/day, it is unlikely that the observed changes in weight were the result of changes in salt and water balance. Thus it seems reasonable to speculate that serum 1,25-(OH)2-D concentrations may vary directly with energy balance, as reflected by changes in body weight. This effect may be mediated by alterations in PO4 metabolism. The accurate assessment of serum 1,25-(OH)2-D levels thus appears to require several measurements over time periods during which body weight is stable.